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Although standardized sediment toxicity testing methods have been developed for the amphipod Hyalella azteca, no standardized chronic water-only toxicity testing methods have been established. Furthermore, optimal feeding and water quality conditions for culturing and toxicity testing with this species remained unclear. The objective of the present study was to determine the food or combination of foods that best promotes survival, growth, and reproduction of the US Lab strain of Hyalella azteca under 42-d, water-only, static-renewal testing conditions. The authors conducted 7 42-d control (no toxicant) tests with various combinations of food (including Tetramin, yeast-cereal leaves-trout chow, diatoms, wheatgrass, alfalfa, and maple leaves) and substrate types (clean "unconditioned" Nitex screens vs "conditioned" Nitex screens that were colonized by live biofilms). Over all treatments, survival ranged from 18% to 96%, dry weight per individual from 0.084 mg to 1.101 mg, and reproduction from 0 young/female to 28.4 young/female. Treatments that included Tetramin tended to result in better performance than those that did not. In particular, treatments that included Tetramin and either conditioned screens or diatoms consistently had high survival, weight, and reproduction values as well as low variability among replicates (measured as coefficient of variation). A ramped Tetramin plus diatom suspension feeding regime appears to have the greatest potential to produce consistently good performance across laboratories using static-renewal systems. Environ Toxicol Chem 2016;35:2407-2415. © 2016 SETAC., (© 2016 SETAC.)